Organelles that illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes

Many anaerobic microbial parasites possess highly modified mitochondria known as mitochondrion-related organelles (MROs). The best-studied of these are the hydrogenosomes of Trichomonas

vaginalis and Spironucleus salmonicida, which produce ATP anaerobically through substrate-level phosphorylation with concomitant hydrogen production; and the mitosomes of Giardia

intestinalis, which are functionally reduced and lack any role in ATP production. However, to understand the metabolic specializations that these MROs underwent in adaptation to parasitism,

data from their free-living relatives are needed. Here, we present a large-scale comparative transcriptomic study of MROs across a major eukaryotic group, Metamonada, examining

lineage-specific gain and loss of metabolic functions in the MROs of Trichomonas, Giardia, Spironucleus and their free-living relatives. Our analyses uncover a complex history of ATP

production machinery in diplomonads such as Giardia, and their closest relative, Dysnectes; and a correlation between the glycine cleavage machinery and lifestyles. Our data further suggest

the existence of a previously undescribed biochemical class of MRO that generates hydrogen but is incapable of ATP synthesis.

M.K., M.M.L. and C.W.S. were supported by a grant (MOP-142349) from the Canadian Institutes of Health Research awarded to A.J.R. This work was also supported, in part, by a grant from the

JSPS Strategic Young Researcher Overseas Visits Program (awarded to R.K.), by NSERC Grant 298366-2009 to A.G.B.S., by a Czech Science Foundation grant to I.Č. (project GA14-14105S) and by

grants from the Japan Society for the Promotion of Science (JSPS; nos 15H05606 and 15K14591 awarded to R.K., 23117005 and 15H05231 awarded to T.H., and 23117006 awarded to Y.I.). We thank A.

A. Heiss for his help with Trimastix marina data generation, and N. Ros for her comments on the manuscript.

Michelle M. Leger, Martin Kolisko, Courtney W. Stairs & Laura Eme

Present address: ‡ Present addresses: Institute of Evolutionary Biology, CSIC-Universitat Pompeu Fabra, Passeig Marítim de la Barceloneta, 37–49, 08003 Barcelona, Spain (M.M.L.); Institute

of Parasitology, Biology Centre, Czech Academy of Sciences, Branišovská 1160/31, 370 05 České Budĕjovice, Czech Republic (M.K.); Department of Cell and Molecular Biology, Uppsala University,

Box 596, 751 24 Uppsala, Sweden (C.W.S. and L.E.),

Michelle M. Leger, Martin Kolisko and Ryoma Kamikawa: These authors contributed equally to this work.

Department of Biochemistry and Molecular Biology, Dalhousie University, 5850 College Street, PO Box 15000, Halifax, Nova Scotia, B3H 4R2, Canada

Michelle M. Leger, Martin Kolisko, Courtney W. Stairs, Laura Eme & Andrew J. Roger

Graduate School of Human and Environmental Studies, Graduate School of Global Environmental Studies, Kyoto University, Yoshida-Honmachi, Sakyo-ku, Kyoto, 606-8501, Japan

Graduate School of Life and Environmental Sciences, Center for Computational Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, 305-8572, Ibaraki, Japan

Department of Zoology, Faculty of Science, Charles University, Vinicna 7, 128 44, Prague 2, Czech Republic

Department of Biological Sciences, University of Arkansas, Fayetteville, 72701, Arkansas, USA

Department of Cell and Molecular Biology, Uppsala University, Box 596, Uppsala, , 751 24, Sweden

Japan Agency for Marine-Earth Science and Technology (JAMSTEC), 2-15, Natsushima-cho, Yokosuka, 237-0061, Kanagawa, Japan

Yantai Institute of Coastal Zone Research, Chinese Academy of Sciences, 17 Chunhui Road, Yantai, 264003, Shandong, People’s Republic of China

Center for Computational Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577, Japan

Department of Biology, Dalhousie University, 1355 Oxford Street, PO Box 15000, Halifax, B3H 4R2, Nova Scotia, Canada

M.K., R.K., J.O.A., Y.I., A.G.B.S., T.H. and A.J.R. conceived and designed the experiments; M.K., K.K., I.Č., J.D.S., F.X., A.Y. and Q.Z. performed the experiments; M.M.L., M.K., R.K.,

C.W.S., K.K., L.E. and Y.I. analysed the data; R.K., C.W.S., J.D.S., K.T., Y.I., A.G.B.S., T.H. and A.J.R. contributed materials and/or analysis tools; and M.M.L., M.K., R.K., A.G.B.S. and

A.J.R. wrote the paper.

Supplementary Figures 1–20, Supplementary Table 1 (PDF 1444 kb)

Predicted mitochondrion-related organelle proteins, and selected predicted cytosolic proteins, in metamonads. (XLSX 149 kb)

File containing the raw phylogenetic trees depicted in Supplementary Figs 4–20, including bootstrap support values, in Newick format. (TXT 539 kb)

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